1. Field of the Invention
The present invention relates to a method for inhibiting NAD(P)H oxidase enzymes in producing reactive oxygen species for growth regulation of normal and malignant tissues.
2. Description of Related Art
Reactive oxygen species (ROS) generated by an NAD(P)H oxidase are important signaling molecules for proliferation of normal cells. The role of a signaling NAD(P)H oxidase has been most extensively explored in vascular smooth muscle cells, where both p22phox and the unique gp91phox homolog NOX1 are important for function of an NAD(P)H oxidase activity that mediates angiotensin II-induced superoxide (O2) formation and redox-sensitive signaling pathways (K. K. Griendling et al. (2000) Circ. Res. 86:494–501; B. Lassegue et al. (2001) Circ. Res. 88:888–894). Similar but structurally distinct NAD(P)H oxidases also perform signaling functions in normal vascular endothelial cells (A. Gorlach et al. (2000) Circ. Res. 87:26–32; S. A. Jones et al. (1996) Am. J. Physiol. 271 (Heart Circ Physiol 49): H1626–H1634) and adventitial cells (P. J. Pagano et al. (1997) Proc. Natl. Acad. Sci. USA (1997) 94:14483–14488), and the gp91phox homolog NOX4 has been described in renal tubular epithelium (M. Geiszt et al. (2000) Proc. Natl. Acad. Sci. USA 97:8010–8014; A. Shiose et al. (2001) J. Biol. Chem. 276:1417–1423), fetal tissue (G. Cheng et al. (2001) Gene 269:131–140), placenta (G. Cheng et al., supra.), and proliferating vascular smooth muscle (13. Lassegue, et al. supra.).
Like normal cells, human tumor cells also produce substantial amounts of ROS spontaneously (B. Del Bello et al. (1999) FASEB J., 13:69–79; D. J. Morre et al. (1995) Proc. Natl. Acad. Sci. USA, 92:1831–1835; T. P. Szatrowski et al. (1991) Cancer Res. 51:794–798), and evidence points to a role for these ROS in signaling neoplastic proliferation. Mitogenic signaling through both Ras (K. Irani et al. (1997) Science 275:1649–1652) and Rac (T. Joneson et al. (1998) J. Biol. Chem. 273:17991–17994) is mediated by O2−, and transfection with mitogenic oxidase NOX1 transforms normal fibroblasts (Y-A. Suh et al. (1999) Nature 410:79–82) and creates cell lines that are tumorigenic in athymic mice (R. Arnold et al. (2001) Proc. Natl. Acad. Sci. USA 98:5550–5555). The NOX1 homolog has been found expressed in the CaCo human colon carcinoma cells (G. Cheng et al., supra; H. Kikuchi et al., (2000) Gene 254:237–243; Y-A. Suh et al, supra) and HepG2 hepatoma cells (H. Kikuchi et al., supra), and gp91phox expression has been demonstrated in small cell lung cancer (D. Wang et al., (1996) Proc. Natl. Acad. Sci. USA 93:13182–13187). However, a potential role for a phagocyte-like NAD(P)H membrane oxidase in signaling proliferation of malignant melanoma cells has not been previously demonstrated.
S. S. Brar and co-workers reported that endogenously produced ROS signal constitutive activation of NF-κB and cellular proliferation in M1619 malignant melanoma cells. Based upon inhibition of these events by the NAD(P)H:quinone oxidoreductase (NQO) inhibitor dicumarol, S. S. Brar and co-workers speculated that cytosolic NQO might provide the enzymatic source of electrons for reduction of membrane ubiquinone to ubiquinol, with subsequent generation of superoxide (O2−) from molecular oxygen. However, dicumarol also inhibited growth of H596 non-small-cell lung cancer cells (S. S. Brar et al. (2001) Am. J. Physiol. Cell Physiol. 280:C659–C676). H596 cells express a mutant NQO protein and have elevated mRNA for NQO but no detectable enzymatic activity (R. D. Traver et al. (1997) Brit. J. Cancer 75:69–75).